Progress in the analysis of membrane protein structure and function

Structural information on membrane proteins is sparse, yet they represent an important class of proteins that is encoded by about 30% of all genes. Progress has primarily been achieved with bacterial proteins, but efforts to solve the structure of eukaryotic membrane proteins are also increasing. Most of the structures currently available have been obtained by exploiting the power of X-ray crystallography. Recent results, however, have demonstrated the accuracy of electron crystallography and the imaging power of the atomic force microscope. These instruments allow membrane proteins to be studied while embedded in the bi-layer, and thus in a functional state. The low signal-to-noise ratio of cryo-electron microscopy is overcome by crystallizing membrane proteins in a two-dimensional protein-lipid membrane, allowing its atomic structure to be determined. In contrast, the high signal-to-noise ratio of atomic force microscopy allows individual protein surfaces to be imaged at subnanometer resolution, and their conformational states to be sampled. This review summarizes the steps in membrane protein structure determination and illuminates recent progress. (C) 2002 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies

Explosive expansion of βγ-Crystallin genes in the ancestral vertebrate

In jawed vertebrates, βγ-crystallins are restricted to the eye lens and thus excellent markers of lens evolution. These βγ-crystallins are four Greek key motifs/two domain proteins, whereas the urochordate βγ-crystallin has a single domain. To trace the origin of the vertebrate βγ-crystallin genes, we searched for homologues in the genomes of a jawless vertebrate (lamprey) and of a cephalochordate (lancelet). The lamprey genome contains orthologs of the gnathostome βB1-, βA2- and γN-crystallin genes and a single domain γN-crystallin-like gene. It contains at least two γ-crystallin genes, but lacks the gnathostome γS-crystallin gene. The genome also encodes a non-lenticular protein containing βγ-crystallin motifs, AIM1, also found in gnathostomes but not detectable in the uro- or cephalochordate genome. The four cephalochordate βγ-crystallin genes found encode two-domain proteins. Unlike the vertebrate βγ-crystallins but like the urochordate βγ-crystallin, three of the predicted proteins contain calcium-binding sites. In the cephalochordate βγ-crystallin genes, the introns are located within motif-encoding region, while in the urochordate and in the vertebrate βγ-crystallin genes the introns are between motif- and/or domain encoding regions. Coincident with the evolution of the vertebrate lens an ancestral urochordate type βγ-crystallin gene rapidly expanded and diverged in the ancestral vertebrate before the cyclostomes/gnathostomes split. The β- and γN-crystallin genes were maintained in subsequent evolution, and, given the selection pressure imposed by accurate vision, must be essential for lens function. The γ-crystallin genes show lineage specific expansion and contraction, presumably in adaptation to the demands on vision resulting from (changes in) lifestyle

Towards nanomicrobiology using atomic force microscopy

At the cross-roads of nanoscience and microbiology, the nanoscale analysis of microbial cells using atomic force microscopy (AFM) is an exciting, rapidly evolving research field. Over the past decade, there has been tremendous progress in our use of AFM to observe membrane proteins and live cells at high resolution. Remarkable advances have also been made in applying force spectroscopy to manipulate single membrane proteins, to map surface properties and receptor sites on cells and to measure cellular interactions at the single-cell and single-molecule levels. In addition, recent developments in cantilever nanosensors have opened up new avenues for the label-free detection of microorganisms and bioanalytes